The present invention generally relates to the field of imaging. In particular, the present invention provides scanning systems and methods for high speed imaging of a sample containing labeled materials, and particularly for scanning arrays of polymer sequences, e.g., oligonucleotide arrays.
Methods and systems for imaging samples containing labeled markers such as confocal microscopes are commercially available. Confocal microscopes generally employ a pinhole that is confocal with the illuminated spot on the specimen to reject light that is not reflected or emitted from objects in the focal plane. This rejection of out-of-focus light enables the microscope to collect and combine a series of optical slices at different focus positions to generate a two or three dimensional representation of the specimen.
Some scanning microscopes employ radiation direction systems, such as galvanometers that include servo-mounted mirrors, to rapidly scan a laser spot across a substrate. Although these microscopes have relatively high scan rates (e.g., on the order of about 30 lines/second or greater), they generally do not achieve both the resolution and field of view that is necessary for some applications, such as imaging an array of sequenced materials on a substrate. In fact, a galvanometer-based confocal microscope's field of view is generally proportional to its resolution. For example, a typical 40x microscope objective, which has a 0.25 .mu.m resolution, has a field size of only about 500 .mu.m. Thus, conventional galvanometer-based confocal microscopes are inadequate for applications requiring both high resolution and a large field of view.
Scanning confocal microscope systems, such as those discussed in U.S. Pat. No. 5,143,854 (Pirrung et al.), PCT WO 92/10092, and U.S. patent application Ser. No. 08/195,889 (now U.S. Pat. No. 5,631,734), incorporated herein by reference for all purposes, are also known. These scanning systems include an optical train which directs a monochromatic or polychromatic light source to about a 5 micron (.mu.m) diameter spot at its focal plane. In some cases, a photon counter detects the emission from the device in response to the light. The data collected by the photon counter represents one pixel or data point of the image. Thereafter, the light scans another pixel as a translation stage moves the device to a subsequent position.
As disclosed, these scanning confocal microscope systems provide high resolution by using an appropriate objective lens and large field of view by using appropriate translation stages. These translation stage-based confocal microscopes, however, obtain high resolution and field of view by sacrificing system throughput. As an example, an array of sequenced material using the pioneering fabrication techniques, such as those disclosed in U.S. Pat. No. 5,143,854 (Pirrung et al.) and U.S. patent application Ser. No. 08/143,312, incorporated herein by reference for all purposes, may have a density of about 10.sup.5 sequences. Assuming that 36 pixels are required for each sequence, the image can take over at least 10 minutes to acquire.
From the above, it is apparent that improved methods and systems for imaging a sample are desired.